Protein phosphatase inhibitory activity of tautomycin photoaffinity probes evaluated at femto-molar level

Herein we describe the further improvement of our in-house developed firefly bioluminescence assay system for the determination of inhibition of protein phosphatase (PP). The advantage with the new system is higher sensitivity as well as being time and sample efficient. The inhibition activity of tautomycin with PP1γ was determined using the upgraded test system and Ki was found to be 4.5 nM, which compare favorably with the activity reported previously by others using different methods. The test system was then used in order to determine the activity of nine tautomycin (TTM) photoaffinity probes. One of the TTM photoaffinity probes (anti-10) was found to possess higher activity than the natural product itself with a Ki of 3.4 nM, while the remaining photoaffinity probes were found to possess Ki in the range of 8.0–213 nM.

Development of a highly sensitive firefly bioluminescence assay system was, in fact, applied to ten tautomycin analogs having photoaffinity probes to obtain the Ki values in the range of 3.4–213 nM, and one of them showed slightly higher activity than the natural tautomycin diacid derivative.Figure optionsDownload as PowerPoint slide